DESCRIPTION (applicant's description): Changes in mitochondrial morphology and copy number are associated with a variety of human diseases including neurological disorders and some types of cancer. Although the metabolic functions of mitochondria have been extensively studied, the molecular mechanisms that regulate mitochondrial membrane dynamics are not understood. In the current funding period, we characterized two different GTPases in S. cerevisiae that regulate mitochondrial morphology and copy number. Dnm1p is a dynamin-related GTPase that acts on the outer mitochondrial membrane to regulate fission. Fzo1p (fuzzy onions) is a transmembrane GTPase that regulates mitochondrial docking and/or fusion. Both GTPases have human homologues that, when mutated, cause defects in mitochondrial morphology. These findings illustrate how effectively yeast can be used as a tool to study the molecular mechanisms that control mitochondrial dynamics in human cells. During the next funding period, we will continue to study mitochondrial fission and fusion in yeast using a combination of genetic, molecular and biochemical approaches. Specifically, we propose: 1) to identify and characterize SFZ genes/proteins required for mitochondrial fission, 2) to determine the role of Fzo1p self-interactions in mitochondrial fusion, 3) to screen for binding partners that interact with Fzo1p in its GTP-bound state, and 4) to develop an in vitro assay for mitochondrial fusion. The studies we propose will provide new information about the molecular and biochemical basis of mitochondrial fission and fusion in eukaryotic cells.